Bioluminescence refers to the production of light by a chemical reaction that originates in a living organism. Primarily a marine phenomenon, bioluminescence is observed in a broad variety of organisms ranging from bacteria to fish, crustaceans and squid, but is primarily attributable to chemical reactions that occur in bacteria and dinoflagellates. Among land dwelling organisms, bioluminescence is found among certain centipedes, millipedes and insects such as fireflies and click beetles but is unknown in plants, birds, reptiles, amphibians and mammals.
Genes responsible for the production of bioluminescent proteins have been isolated by molecular cloning techniques, leading to the widespread and expanding use of such proteins in a variety of applications including, inter alia, use as: 1) bioreporters for detection of environmental compounds, 2) reagentless components in bioelectronic devices, 3) whole cell logic gates for biocomputing, 4) tools for in situ functional imaging, 5) markers of recombinant strains released into the environment, and 6) markers to enable in vivo imaging of bacteria during systemic infection in animal hosts (Applegate et al., Appl. Environ. Microbiol., 64:2730-2735, 1998; Sayler, G. S., and Ripp, S., Current Opinion in Biotechnology, 11:286-289, 2000; and Liu et al., Plasmid, 44:250-261, 2000).
Luciferases of the lux family (Lux) are bioluminescent proteins derived from certain bioluminescent bacterial strains that have found widespread use in various applications. Lux is the product of the luxAB genes, which encode luciferase, a heterodimeric bioluminescent catalyzing enzyme. The entire luxCDABE cassette has been isolated from several bioluminescent bacteria including Vibrio fischeri, V. harveyi, and Photorhabdus luminescens. Use of the complete operon of bacterial luciferase (i.e., luxCDABE) has enabled construction of self-sufficient bioreporters containing all of the required elements for production of light in response to specific chemicals or physical agents.
Despite these considerable advances, currently available bioreporters based on luxCDABE are not ideally suited for applications requiring a relatively short half life of the reporter protein. As one example, the duration of Lux emission by existing bioreporters limits their usefulness for monitoring certain events in real time, such as both initiation and termination of transcription in a cell. Additionally, in applications using bioreporters in synthetic gene circuits, the long time constants associated with decay of presently available reporter proteins can significantly degrade circuit performance.